Summary
There is a wealth of information about monoclonal antibody (MAb) specificity and function on fixed tissues, yet little is known about formation and release of antigen-antibody complexes and their functional behavior in vivo.
We analyzed the pathway of radiolabeled MAbs directed against melanoma-associated antigens by radioimmunoelectron microscopy (RIEM) on metabolically active cells of the melanoma cell lines SK-MEL-28, MeWo and Colo 38 at different time intervals. In parallel, binding and release of MAbs were investigated by the radioantibody binding assay (RBA). Both procedures gave essentially concordant results.
Preferentially stable binding of immune complexes (ICs) to the cell surface after 30 and 120 min was shown for the MAb L10. Internalization was demonstrated for the MAb M.2.9.4. At the ultrastructural level, direct evidence of this phenomenon was obtained by visualization of radioactivity within the cytoplasm after 120 min. In the RBA this process was indicated by resistance of bound MAbs to acid buffer desorption. RIEM pointed to different transport mechanisms: constitutive internalization by endocytotic vesicles, or receptor-mediated endocytosis by coated vesicles. Shedding was indicated for the MAb R24 by release of the ICs from the cell membrane. It was demonstrated that stable fixation of ICs on the cell surface or modulation by internalization led to high accumulation rates, while shedding of antigen-antibody complexes resulted in a low accumulation of the MAb in tumor cells.
Assuming that the potential of MAbs for clinical application is determined by the biological behavior of antigen-antibody complexes, these methods are suitable for demonstration of antigenic modulation by MAbs and eventually enable us to predict the localization, penetration and distribution pattern of individual MAbs in the melanoma patient.
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References
Alberts B, Bray D, Lewis J, Raff M, Roberts K, Watson JD (eds) (1983) Molecular biology of the cell. Garland, New York
Bouteille M (1976) The “LIGOP” method for routine ultrastructural autoradiography. J Microsc Biol Cell 27:121–128
Bröcker EB, Suter L, Brüggen J, Ruiter DJ, Macher E, Sorg C (1985) Phenotypic dynamics of tumor progression in human malignant melanoma. Int J Cancer 36:29–35
Brown JP, Nishiyama K, Hellström I, Hellström KE (1981) Structural characterization of human melanoma-associated antigen p97 with monoclonal antibodies. J Immunol 127:539–546
Brüggen J, Sorg C (1983) Detection of phenotypic differences on human malignant melanoma lines and their variant sublines with monoclonal antibodies. Cancer Immunol Immunother 15:200–205
Carrasquillo JA, Krohn KA, Beaumier P, McGuffin RW, Brown JP, Hellström KE, Hellström I, Larson SM (1984) Diagnosis of and therapy for solid tumors with radiolabeled antibodies and immune fragments. Cancer Treat Rep 68:317–328
Casellas P, Brown JP, Gros O, Gros P, Hellström I, Jansen FK, Poncelet P, Roncucci R, Vidal H, Hellström KE (1982) Human melanoma cells can be killed in vitro by an immunotoxin specific for melanoma-associated antigen p97. Int J Cancer 30:437–443
Chatenoud L, Bach J-F (1984) Antigenic modulation — a major mechanism of antibody action. Immunol Today 5:20–25
Dautry-Varsat A, Lodish HF (1984) Zellrezeptoren als Transportsystem. Spektr Wissensch 7:98–105
Dippold WG, Lloyd KO, Li LTC, Ikeda H, Oettgen HF, Old LJ (1980) Cell surface antigens of human malignant melanoma: definition of six antigenic systems with mouse monoclonal antibodies. Proc Natl Acad Sci USA 77:6114–6118
Dippold WG, Knuth A, Meyer zum Büschenfelde K-H (1984) Inhibition of human melanoma cell growth in vitro by monoclonal anti-GD3-ganglioside antibody. Cancer Res 44:806–810
Dippold WG, Knuth KRA, Meyer zum Büschenfelde K-H (1985) Inflammatory tumor response to monoclonal antibody infusion. Eur J Cancer Clin Oncol 21:907–912
Garrigues HJ, Tilgen W, Hellström I, Franke W, Hellström KE (1982) Detection of a human melanoma-associated antigen, p97, in histological sections of primary human melanomas. Int J Cancer 29:511–515
Hellström I, Brown JP, Hellström KE (1983) Melanoma-associated antigen p97 continues to be expressed after prolonged exposure of cells to specific antibody. Int J Cancer 31:553–555
Hellström KE, Hellström I, Brown JP (1984) Diagnostic and therapeutic use of monoclonal antibodies to human tumor antigens. Med Oncol Tumor Pharmacother 1:143–147
Houghton AN, Eisinger M, Albino AP, Cairncross JG, Old LJ (1982) Surface antigens of melanocytes and melanomas. J Exp Med 156:1755–1766
Houghton AN, Mintzer D, Cordon-Cardo C, Welt S, Fliegel B, Vadhan S, Carswell E, Melamed MR, Oettgen HF, Old LJ (1985) Mouse monoclonal IgG3 antibody detecting GD3 ganglioside: a phase I trial in patients with malignant melanoma. Proc Natl Acad Sci USA 82:1242–1246
Köhler G, Milstein C (1975) Continuous cultures of fused cells secreting antibody of predefined specificity. Nature 256:495–497
Knuth A (1986) Response to monoclonal antibodies in melanoma: specific or nonspecific? Eur J Cancer Clin Oncol 22:1285–1288
Lloyd KO (1983) Human tumor antigens: detection and characterization with monoclonal antibodies. In: Herbermann R (ed) Basic and clinical tumor immunology, vol 1. Martinus Nijhoff, The Hague, pp 159–214
Matzku S, Tilgen W (1985) Experimentelle Grundlagen der Radioimmundiagnostik und-therapie humaner Melanome mit monoklonalen Antikörpern. In: Holzmann H, Altmeyer P, Hör G, Hahn K (eds) Dermatologie und Nuklearmedizin. Springer, Berlin Heidelberg New York Tokyo, pp 134–147
Matzku S, Mattern J, Georgi P, Hellström I (1982) Radioimmunolocalization of melanomas with monoclonal antibodies directed against the p97 marker. Model experiments in vitro and in nude mice bearing transplanted human melanomas. In: Höfer R, Bergmann H (eds) Radioaktive Isotope in Klinik und Forschung, Bd 15. Egermann, Vienna, pp 295–301
Matzku S, Bröcker E-B, Brüggen J, Dippold WG, Tilgen W (1986) Modes of binding and internalization of monoclonal antibodies to human melanoma cell lines. Cancer Res 46:3848–3854
Natali PG, Aguzzi A, Veglia F, Imai K, Burlage RS, Giacomini P, Ferrone S (1983) The impact of monoclonal antibodies on the study of human malignant melanoma. J Cutan Pathol 10:514–528
Ng A-K, Giacomini P, Kantor R, Ferrone S (1982) Molecular heterogeneity and shedding of a high-molecular-mass melanoma-associated antigen identified with monoclonal antibodies. Clin Chem 28:2347–2350
Old LJ, Stockert E, Boyse EA, Kim JH (1968) Antigenic modulation. Loss of TL antigen from cells exposed to TL antibody. Study of the phenomenon in vitro. J Exp Med 127:523–531
Oldham RK, Foon KA, Morgan AC, Woodhouse CS, Schroff RW, Abrams PG, Fer M, Schoenberger CS, Farrell M, Kimball E, Sherwin SA (1984) Monoclonal antibody therapy of malignant melanoma: in vivo localization in cutaneous metastasis after intravenous administration. J Clin Oncol 2:1235–1244
Pastan I, Willingham MC (1983) Receptor-mediated endocytosis: coated pits, receptosomes and the Golgi. Trends Biochem Sci 8:280–254
Pernis B (1985) Internalization of lymphocyte membrane components. Immunol Today 6:45–49
Pesando J, Ritz J, Lazarus H, Tomaselli K, Schlossman S (1981) Fate of a common acute lymphoplastic leukemia antigen during modulation by monoclonal antibody. J Immunol 126:540–544
Real FX, Mattes MJ, Houghton AN, Oettgen HF, Lloyd KO, Old LJ (1984) Class 1 (unique) tumor antigens of human melanoma. J Exp Med 160:1219–1233
Reisfeld RA, Cheresh DA (1985) Human tumour-associated antigens: targets for monoclonal antibody-mediated cancer therapy. Cancer Surveys 4:271–290
Reisfeld RA, Ferrone S (eds) (1982) Melanoma antigens and antibodies. Plenum, New York
Rowland GF, Axton CA, Baldwin RW, Brown JP, Corvalan JRF, Embleton MJ, Gore VA, Hellström I, Hellström KE, Jacobs E, Marsden CH, Pimm MV, Simmonds RG, Smith W (1985) Antitumor properties of vindesine-monoclonal antibody conjugates. Cancer Immunol Immunother 19:1–7
Siccardi AG, Buraggi GL, Callegaro L, Mariani G, Natali PG, Abbati A, Bestagno M, Caputo V, Mansi L, Masi R, Paganelli G, Riva P, Salvatore M, Sanguineti M, Troncone L, Turco GL, Scassellati GA, Ferrone S (1986) Multicenter study of immunoscintigraphy with radiolabeled monoclonal antibodies in patients with melanoma. Cancer Res 46:4817–4822
Suter L, Bröcker EB, Brüggen J, Ruiter DJ, Sorg C (1983) Heterogeneity of primary and metastatic human malignant melanoma as detected with monoclonal antibodies in cryostat sections of biopsies. Cancer Immunol Immunother 16:53–58
Suter L, Brüggen J, Bröcker EB, Sorg C (1985) A tumor-associated antigen expressed in melanoma cells with lower malignant potential. Int J Cancer 35:787–791
Tilgen W, Hellström I, Engstner M, Garrigues HJ, Riehl R, Hellström KE (1983) Localization of melanoma-associated antigen p97 in cultured human melanoma, as visualized by light and electron microscopy. J Invest Dermatol 80:459–463
Tilgen W, Dzarlieva RT, Breitkreutz D, Hennes B, Engstner M, Matzku S, Fusenig NE (1985) Heterogeneity of human malignant melanoma cells in vivo and in vitro: role of experimental systems. In: Bagnara J, Klaus SN, Paul E, Schartl M (eds) Pigment cell 1985 Biological, molecular and clinical aspects of pigmentation. University of Tokyo Press, Tokyo, pp 435–447
Tilgen W, Dippold W, Matzku S (1986) Immundiagnostik und Immuntherapie maligner Melanome mit monoklonalen Antikörpern: experimentelle und klinische Ergebnisse. In: Rü** K-W, Stary A, Tronnier H (eds) I. Dermatologisches Forum, Dortmund: Neues in der Therapie. Verlag medical concept, Neufahrn, pp 216–224
Vermeer BJ, Wijsman MC, Mommaas-Kienhuis AM, Ponec M (1986) Binding and internalization of low-density lipoproteins in SCC25 cells and SV40 transformed keratinocytes. A morphologic study. J Invest Dermatol 86:195–200
Vitetta ES, Krolick KA, Miyama-Inaba M, Cushley W, Uhr JW (1983) Immunotoxin: a new approach to cancer therapy. Science 219:644–650
Wolff HH (1976) Elektronenmikroskopische Autoradiographie. Hautarzt 27 [Suppl I]: 198–201
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This work was supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 136 “Krebsforschung”
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Tilgen, W., Matzku, S., Kaufmann, I. et al. Modulation of melanoma-associated antigens by monoclonal antibodies as visualized by radioimmunoelectron microscopy and radioantibody binding assay. Arch Dermatol Res 279 (Suppl 1), S116–S126 (1987). https://doi.org/10.1007/BF00585934
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DOI: https://doi.org/10.1007/BF00585934